1. Field of the Invention
The present invention generally relates to compounds having a scytoneman skeleton, such as scytonemin. Specifically, the present invention relates to methods of using a compound having a scytoneman skeleton to inhibit, prevent, modulate, or attenuate cell-cycle progression, cell proliferation, kinase activity, phosphorylation, or induce apoptosis. Additionally, the present invention relates to methods of treating, preventing, or inhibiting diseases and disorders associated with cell cycle progression, cell proliferation, kinase activity, or angiogenesis in a subject comprising administering a scytoneman compound to the subject.
2. Description of the Related Art
The cell cycle is a tightly regulated series of events, eventually leading to the formation of two daughter cells. The transitions of the cell cycle are controlled by the reversible phosphorylation of certain proteins. For example, the cyclin-dependent kinases (CDKs) when complexed with a specific cyclin protein, are responsible for coordinating the passage of the cell through the different stages of the cell cycle. See Norbury, C. et al. (1992) Ann. Rev. Biochem. 61:441-470; Sherr, C. J. (1993) Cell 73:1059-1065; and Nasmyth, K. (1993) Curr. Opin. Cell Biol. 5:166-179. The phosphorylation of certain residues on CDK/cyclin complexes control catalytic activity both positively or negatively. See De Bondt, H. L. et al. (1993) Nature 363:595-602; Ducommun, B. et al. (1991) EMBO J 10:3311-3319; Jeffrey, P. D. et al. (1995) Nature 376:313-320. For instance, the catalytic activity of CDK1/cyclin B requires the removal of inhibitory phosphates by the cdc25C phosphatase. See Dunphy, W. G. et al. (1991) Cell 67:189-196. Subsequent activity of CDK1/cyclin B allows the cycle to progress from G2 into mitosis. See Maller, J. L. (1991) Curr. Opin. Cell Biol. 3:269-275; Krek, W. et al. (1991) EMBO J 10:3321-3329; Atherton, F. S. et al. (1993) Mol. and Cell. Biol. 13:1675-1685; Endicott, J. A. et al. (1994) Prot. Eng. 7:243-253.
Aberrant cell proliferation is symptomatic of many debilitating disease conditions including rheumatoid arthritis, chronic obstructive pulmonary disorder, psoriasis, and cancers. Genetics research has advanced our knowledge as to what factors contribute to regulating the cell cycle. Many of these molecules have been targeted for pharmacological inactivation in attempts to discover novel mechanisms by which uncontrolled cell proliferation may be abrogated.
Recent evidence indicates that a novel family of enzymes, the polo-like kinases (Plks), may serve to regulate the activity of cdc25C. A Xenopus Plk (Plx1), and the human Plk homologue (PKL1), have both been implicated in the phosphorylation and subsequent activation of cdc25C. See Kumagai, A. et al. (1996) Science 273:1377-1380; Roshak, A. K. et al. (2000) Cellular Signaling 12:405-411. These findings suggest that the Plks constitute another level of regulation for CDKs, in particularly the activation of the CDK1/cyclin B complex via activation of cdc25C.
The polo gene was originally identified in Drosophila melanogaster, where mutant polo phenotypes displayed aberrant mitotic divisions due to abnormal spindle formation. See Sunkel and Glover, (1988) J. Cell Sci. 89:25-38. Functional homologues have subsequently been identified in organisms ranging from protozoa to human. See Graham, T. M. et al. (1998) Gene 207:71-77; Kitada, K. et al. (1993) Mol. Cell. Biol. 13:4445-4457; Ohkura et al. (1995) Genes and Dev. 9:1059-1073; Kumagai (1996); Clay et al. (1993) PNAS USA 90: 4882-4886; Golsteyn, et al. (1994) J. Cell Sci. 107:1509-1517; Hamanaka, et al. (1994) Cell Growth Differ. 5:249-257. Members of this family of serine/threonine kinases share homologous structural features which includes a 30 amino acid motif at the C-terminus, called the polo-box, the function of which is unknown. See Golsteyn, et al (1996) Cell Cycle Res. 2:107-114; and Nigg (1998) Curr. Opin. Cell Biol. 10:776-783. Polo-family members also share a conserved sequence in the ATP binding domain. This conserved sequence, GxGGxAxC, differs from the sequences of ATP binding pockets of most other serine/threonine kinases, GxGxxGxV, wherein xe2x80x9cxxe2x80x9d is any nucleotide. See Golsteyn et al. (1996) and Nigg (1998).
Functional studies suggest that Plk homologues play an integral role in regulating cell cycle progression. See Lane, et al. (1996) J. Cell Biol. 135:1701-1713; Llamazares, et al. (1991) Genes Dev. 5:2153-2165; and Ohkura et al. (1995). The expression of PLK1 correlates with the mitotic index of many cancer cell lines. Over-expression of the enzyme in murine NIH3T3 cells results in a transformed phenotype. See Smith, et al. (1997) Biochem. Biophys. Res. Commun. 150:1165-1172. In addition, when PLK1-neutralizing antibodies are microinjected into a T cell line, the T cells lose the ability to undergo mitosis. See Lane (1996). These data suggest that PLK1 plays a critical role in cell cycle regulation and targeting its activity may yield another means of inhibiting aberrant cell proliferation.
Prior to the present invention, there have been no reports of small molecule inhibitors of PLK1 activity. Thus, there is a need for a polo-like kinase inhibitor for the treatment of cancer and other diseases and disorders associated with aberrant cell cycle progression and cell proliferation.
In some embodiments, the present invention relates to a method of inhibiting, modulating, attenuating, or preventing cell-cycle progression, intracellular signaling, or cell proliferation of a cell comprising exposing the cell to at least one scytoneman compound. In preferred embodiments, the scytoneman compound is scytonemin.
In some embodiments, the present invention relates to a method of inhibiting, modulating, attenuating, or preventing a kinase, kinase activity, or phosphorylation of a substrate by a kinase comprising exposing the kinase to at least one scytoneman compound. In preferred embodiments, the kinase is a cell cycle regulatory kinase or an intracellular signaling kinase. In some embodiments, the kinase is a serine/threonine kinase, a threonine/tyrosine kinase, or a tyrosine kinase, preferably, a polo-like kinase, a cyclin-dependent kinase, or a checkpoint kinase, more preferably, PLK1, Myt1, CHK1, CDK1/cyclinB, PKCxcex22, PKA, or Tie2.
In some embodiments, the present invention relates to a method of treating, preventing, or inhibiting a disease or disorder associated with inflammation, cell cycle progression, cell proliferation, kinase activity, tissue hyperplasia or angiogenesis in a subject comprising administering at least one scytoneman compound to the subject. Preferably, the subject is a mammal, more preferably, human. In some embodiments, the disease or disorder is an acute or a chronic inflammatory disease or disorder. In some embodiments, the disease or disorder is cancer, a papilloma, rheumatoid arthritis, psoriasis, atherosclerosis, diabetic retinopathy, chronic obstructive pulmonary disorder, tuberculosis, chronic cholecystitis, osteoarthritis, rheumatic carditis, bronchiectasis, Hashimoto""s thyroiditis, ulcerative colitis, Crohn""s disease, silicosis, or the like. In preferred embodiments, the disease or disorder is cancer, rheumatoid arthritis, angiogenesis, or psoriasis. Preferably, the scytoneman compound is administered in a therapeutically effective amount or concentration.
In some embodiments, the present invention relates to a method of inducing apoptosis in a cell comprising exposing the cell to at least one scytoneman compound. In some embodiments, the present invention relates to a method of treating a disease, disorder, or tissue, in a subject comprising inducing apoptosis in a target by administering at least one scytoneman compound to the subject. In preferred embodiments, the target is a cell, an abnormal cell, or an organism.
In some embodiments, the present invention relates to a method of treating, preventing, inhibiting, attenuating or modulating inflammation related to an neurogenic inflammatory pathway in a subject comprising administering at least one scytoneman compound to the subject.
In some embodiments, the present invention relates to a method of treating, preventing, or inhibiting pain associated with neurogenic inflammation in a subject comprising administering at least one scytoneman compound to the subject.
In some embodiments, the present invention relates to a method of treating, preventing, or inhibiting a tissue hyperplasia in a subject comprising administering to the subject a composition comprising at least one scytoneman compound. In preferred embodiments, the scytoneman compound is scytonemin. Preferably, the composition is topically administered to the subject. In preferred embodiments, the scytoneman compound is present in a therapeutically effective amount. The subject is preferably mammalian, and more preferably, the subject is human. The composition may further include a pharmaceutical excipient. The composition may also include at least one supplementary active compound. Preferably, the supplementary active compound is an analgesic, an anti-inflammatory agent, or an anti-proliferative agent such as dexamethasone, glucocorticoids, cyclooxygenase (COX) inhibitors, aspirin, methotrexate, taxol, and the like. In some preferred embodiments, the tissue hyperplasia is skin hyperplasia. Preferably, the tissue hyperplasia is prevented or inhibited by about 25% to about 75% as compared to a control.
In some embodiments, the present invention relates to a method of reducing an epidermal layer in a subject comprising administering to the subject a composition comprising at least one scytoneman compound. In preferred embodiments, the scytoneman compound is scytonemin. Preferably, the composition is topically administered to the subject. In preferred embodiments, the scytoneman compound is present in a therapeutically effective amount. The subject is preferably mammalian, and more preferably, the subject is human. The composition may further include a pharmaceutical excipient. The composition may also include at least one supplementary active compound. Preferably, the supplementary active compound is an analgesic, an anti-inflammatory agent, or an anti-proliferative agent such as dexamethasone, glucocorticoids, cyclooxygenase (COX) inhibitors, aspirin, methotrexate, taxol, and the like. In preferred embodiments, the epidermal layer is reduced by about 1 to about 6 cells or about 0.02 mm to about 0.04 mm.
In some embodiments, the present invention relates to a method of reducing, preventing, or inhibiting cell proliferation in a subject comprising administering at least one scytoneman compound to the subject. In preferred embodiments, the scytoneman compound is scytonemin. Preferably, the composition is topically administered to the subject. In preferred embodiments, the scytoneman compound is present in a therapeutically effective amount. The subject is preferably mammalian, and more preferably, the subject is human. The composition may further include a pharmaceutical excipient. The composition may also include at least one supplementary active compound. Preferably, the supplementary active compound is an analgesic, an anti-inflammatory agent, or an anti-proliferative agent such as dexamethasone, glucocorticoids, cyclooxygenase (COX) inhibitors, aspirin, methotrexate, taxol, and the like. In preferred embodiments, the cell proliferation is reduced, prevented, or inhibited by about one third as compared to a control.
In some embodiments, the present invention relates to a method of reducing the concentration of at least one proinflammatory mediator in a subject comprising administering to the subject a therapeutically effective amount of a scytoneman compound. In preferred embodiments, the scytoneman compound is scytonemin. Preferably, the composition is topically administered to the subject. In preferred embodiments, the scytoneman compound is present in a therapeutically effective amount. The subject is preferably mammalian, and more preferably, the subject is human. The composition may further include a pharmaceutical excipient. The composition may also include at least one supplementary active compound. Preferably, the supplementary active compound is an analgesic, an anti-inflammatory agent, or an anti-proliferative agent such as dexamethasone, glucocorticoids, cyclooxygenase (COX) inhibitors, aspirin, methotrexate, taxol, and the like. The proinflammatory mediator is IL-1xcex2, TNFxcex1, PGE2, or the like.
In some embodiments, the present invention relates to mouse ear assay for determining whether a test compound has an effect on chronic inflammation, cell proliferation, or tissue hyperplasia comprising measuring the amount of IUdR uptake in a test sample, or histologically analyzing a test sample after staining with PCNA, or both.
The present invention also relates to a method for treating, preventing, or inhibiting an infection, disease, or disorder related to an organism belonging to the kingdom Protista in a subject comprising administering to the subject a therapeutically effective amount of a scytoneman compound. The organism may be a flagellate, a ciliate, an opalinidae, or a sporozoan. The organism may be a plasmodium, a trypanosome, or a paramecium such as trichinosis, trypanosomiasis, leishmania, filariasis, or dracunculiasis. The infection, disease or disorder may be malaria, Chagas"" disease, African sleeping sickness, Leishmaniasis, giardiasis, or amebic dysentery.
It is to be understood that both the foregoing general description and the following detailed description are exemplary and explanatory only and are intended to provide further explanation of the invention as claimed. The accompanying drawings are included to provide a further understanding of the invention and are incorporated in and constitute part of this specification, illustrate several embodiments of the invention and together with the description serve to explain the principles of the invention.